Group A Streptococcus (GAS) is an exclusive human pathogen that causes significant morbidity and mortality. Given the recent rise in GAS invasive disease outbreaks, antibiotic resistance in GAS and lack of a human vaccine, the need to identify novel antimicrobial targets to improve infection control cannot be ignored. The bacterial intercellular communication machinery offers a promising target for antibacterial drug discovery as it provides a means of infection control other than growth inhibition. GAS global transcription regulator, RopB, controls the expression of several virulence factors including secreted protease, SpeB, which is required for host tissue damage and disease dissemination. Previous data suggested that RopB requires growth-phase-specific environmental signals to mediate gene regulation. Recently, through the collaborative effort of laboratories with expertise in structural biology, biophysics, bacterial genetics, and bacterial pathogenesis, we demonstrated that RopB is a component of GAS communication machinery and it uses growth phase-specific secreted peptides as intercellular signals to coordinate virulence gene regulation. Although we identified the secretion signal sequence of Vfr as RopB inhibition signal at low cell density, the genetic identity of activation signals specific for high cell density remains unknown. This research proposal is designed to identify and characterize the peptide signals and assess their impact on GAS virulence. The objective of this proposal is to enhance molecular understanding of the intercellular signaling machinery in GAS with the long-term goal of devising anti-infective strategies that target bacterial communication. Successful completion of the proposed study would not only provide the molecular and mechanistic details of the intercellular signaling in -hemolytic streptococci but also could potentially provide the scaffold for the development of future anti-infective drugs.